While the concept of beta cell replacement as a therapy for diabetes seems straightforward, a major obstacle for this therapy has been the limited amount of available islet tissue for transplantation. 1 approach has been to manipulate adult human pancreatic tissue in vitro. There are 2 general results obtained from these experiments: 1) from islet depleted tissue limited expansion but with increased numbers of functional beta cells and 2) from islet enriched preparations extensive expansion with multiple passages with low levels of islet markers after further manipulation. A rigorous demonstration of pancreatic progenitor cells from the adult human pancreas has not yet been shown. It would be immaterial whether such cells were a product of culture and did not exist in vivo if in vitro progenitors could be identified, expanded and differentiated to provide a reliable source of functional beta cells. We propose to develop an in vitro genetic lineage tracing system based on a reporter lenti virus that through the Cre-lox system can specifically mark a particular initial cell type and all its progeny. A system to do genetic lineage tracing in human tissue in vitro would be an important tool in the quest for differentiating beta cells as well as for any other in vitro differentiation model. By genetically marking different pancreatic cell types and then following through the protocol for in vitro generation of new insulin-producing cells, we should be able to identify the original cell or cell types that give rise in vitro to the insulin-producing cells. Identifying such cell(s) would then direct focus on the expansion and differentiation of that cell type to a sustainable source of new beta cells. [unreadable] [unreadable]